Project 2 - The role of Foxn1 in controlling the transition from thymus expansion to homeostasis

摘要 越来越多的证据表明，胸腺上皮细胞(TEC)是负责协调 胎儿发育及产后功能。一个单一的转录因子FOXN1已知是一个关键的 在整个生命周期中，TEC的增殖和功能的多个方面的调节器。TEC车厢 从胎儿/新生儿扩张计划切换到青少年动态平衡计划，有一个关键的转折点 在小鼠出生后约7天发生(P07)，在人类约4个月大时发生。在老鼠身上的数据表明 这种转换是由视网膜母细胞瘤(RB)途径控制的，至少部分是通过抑制Foxn1基因发挥作用 由E2F转录因子表达。缺乏多个Rb家族成员的小鼠无法进行这种转换 并继续扩张。K5.D_1转基因株系，其中细胞周期蛋白D_1在TEC中特异性高表达， 通过激活抑制Rb功能的细胞周期蛋白依赖的激酶来模拟这种Rb功能丧失的表型。在这两个地方 Rb突变体和K5.D1转基因株Foxn1基因表达上调，并抑制Foxn1基因表达 使用出生后特定的亚型等位基因(Foxn1Z/Z)可以恢复胎儿到幼年的转换，使胸腺正常化 尺码。我们还表明，Foxn1对TEC增殖的调节主要发生在MHCIIlo TEC中，一致 根据Richie实验室的初步数据，Sca1-MHCIIlo TEC子集可能包含一个关键的增殖 在从胎儿扩张到幼年动态平衡的转换中，祖细胞种群。这些数据表明 在围产期到幼年期的转变过程中，Rb蛋白通过E2F转录因子调节Foxn1的转录 在特定的TEC亚群中，不同的TEC可以“刹车”TEC的增殖，从而调节器官的大小。那里 也有越来越多的证据表明，新生儿和成人胸腺具有阶段特定的功能，可以产生不同的T细胞 细胞群。由于FOXN1也被认为是TEC分化的关键调节因子，Foxn1如何变化 调控TEC的表达以控制TEC的增殖也应对TEC的功能产生重大影响 新生儿和成人胸腺。我们提出了三个具体目标来检验这一假设，即RB依赖 新生儿期Foxn1基因表达水平的变化与器官建立的关系 动态平衡也需要产生特定的微环境，这是新生儿专一性所必需的 器官功能。该项目的目标是确定：Foxn1的表达在整个 过渡影响小鼠胸腺间质成分的变化，并与人胸腺的间质变化进行比较； 在这一转变过程中，Foxn1的表达如何受到Rb通路的调节；以及这些Foxn1依赖于 过程会影响免疫功能。

Abstract There is growing evidence that thymic epithelial cells (TEC) are the key cell type responsible for orchestrating the fetal development and postnatal function. A single transcription factor, FOXN1, is known to be a critical regulator of multiple aspects of TEC proliferation and function throughout the lifespan. The TEC compartment switches from a fetal/neonatal expansion program to a juvenile homeostasis program, with a key transition point occurring at about 7 days postnatal in mice (P07), and about 4 months of age in humans. Data in mice show that this switch is controlled by the retinoblastoma (RB) pathway, acting at least in part by suppression of Foxn1 gene expression by E2F transcription factors. Mice deficient for multiple RB family members fail to make this switch and continue to expand. The K5.D1 transgenic line, in which CyclinD1 is specifically overexpressed in TEC, mimics this RB loss of function phenotype by activating cyclin-dependent kinases that inhibit RB function. In both RB mutants and K5.D1 transgenics Foxn1 gene expression is elevated, and suppressing Foxn1 expression using a postnatal-specific hypomorphic allele (Foxn1Z/Z) restores the fetal to juvenile switch, normalizing thymus size. We have also shown that Foxn1 regulation of TEC proliferation occurs primarily in MHCIIlo TEC, consistent with preliminary data from the Richie lab that a Sca1-MHCIIlo TEC subset may contain a key proliferating progenitor population during the switch from fetal expansion to juvenile homeostasis. These data suggest that during the perinatal to juvenile transition, RB proteins modulate Foxn1 transcription via E2F transcription factors differentially in specific TEC subsets to ‘put the brakes on’ TEC proliferation, and thus regulate organ size. There is also growing evidence that the neonatal and adult thymi have stage-specific functions that generate distinct T cell populations. As FOXN1 is also known to be a key regulator of TEC differentiation, the changes in how Foxn1 expression is regulated to control TEC proliferation should also have significant impacts on TEC function in the neonatal and adult thymus. We propose three specific aims to test the hypothesis that the RB-dependent changes in Foxn1 gene expression levels during the neonatal period that are essential for establishing organ homeostasis are also required to generate specific microenvironments that are necessary for neonatal-specific organ functions. The goals of this project are to determine: how changes in Foxn1 expression across this transition impact stromal composition changes in mouse, and compare to stromal changes in human thymus; how Foxn1 expression is regulated by the RB pathway during this transition; and how these Foxn1-dependent processes impact immune function.